Not applicable.
1. Field of the Invention
This invention relates to methods for increasing resistance of plants to plant pathogens. More specifically, this invention relates to the surprising discovery that the application to plants of one or more reactive oxygen species and of one or more plant systemic inducers, either simultaneously or within a short time of each other, results in an increase in the level of pathogenesis-related proteins and of systemic acquired resistance in the plants over the effect of either one alone.
2. Background
Commercial cultivation of plants is a major part of the economy, encompassing not only crops grown for human food and animal feed, but also those, like cotton, grown for fiber, and others, such as flowers, grown for beauty. The importance of plants to people and to the economy can hardly be overstated. Plants are, however, also subject to constant attack by insects, fungi, bacteria, viruses, nematodes, and other pathogens. When pathogens find susceptible plants, these attacks can result in the loss of yield and quality, and may result in the loss of entire crops. These losses result in substantial economic harm to the growers and, in some areas of the world, contribute to famine.
Except for those farmers who practice organic farming, most attempts to control pathogens involve the use of pesticides, such as fungicides and insecticides. Many pesticides, however, have been withdrawn from the market because they have undesirable environmental impacts, and many currently on the market are being scrutinized for their environmental impact and may be withdrawn in the future. In addition, few, if any, pesticides are effective against the full range of pests which may attack a given crop from sowing to harvest to post-harvest storage. Thus, a number of different pesticides with different target organisms may need to be applied. Each one must be applied at the correct time in the growth of the plants to provide effective control of the target organism, each has its own requirements for handling and application, and each may require different, specialized equipment. Moreover, many pesticides are toxic or have toxic residues, and their use is therefore often restricted to certain windows of time before harvest, after which they cannot be used because of the potential danger to the consumer. During this window, the crop may be essentially unprotected, or yet another agent, safer for use close to harvesting, may be needed. The use of traditional chemical agents therefore requires complicated planning, careful timing, and considerable effort.
While pesticides form the bulk of attempts by farmers to protect plants from pathogenic attack, not all protection of plants against pathogens comes from the application of pesticides. For decades, it has been known that plants also have a wide variety of structural and biochemical defenses against attack by pathogens. See, e.g., Agrios, G., Plant Pathology, Academic Press, San Diego Calif. (3rd ed., 1988).
One of the biochemical defenses produced by plants in response to attack is induced resistance, in which plants which have been inoculated with biological agents or pretreated with various chemicals develop nonspecific resistance not only to the initial agent itself, but also to a variety of pathogenic agents, such as viruses, fungi, bacteria, and some insects. Induced resistance usually commences in the area around the initial inoculation, but over the course of a few days, may spread to portions of the plant not inoculated, a phenomenon known as systemic induced resistance, or as systemic acquired resistance (xe2x80x9cSARxe2x80x9d).
A number of compounds, such as salicylic acid, can induce resistance. See, e.g., Klessig, D. and Malamy, J., Plant Mol. Biol., 26:1439-1458 (1994); Raskin, I., Annu. Rev. Plant Physiol. Plant Mol. Biol. 43:439-463 (1992). They can be used to induce local resistance, by injection or spraying, or to induce SAR when absorbed, for example, through the roots. See generally, Agrios, supra, at Chapters 5 and 9. SAR develops some 7 days or more after exposure to the inoculant or chemical agent, and usually lasts for some 3 to 5 weeks. Id.
Because SAR protects plants against many different pests, increasing SAR in crops could potentially decrease or even eliminate the need to apply toxic pesticides. Further, since SAR protects against a multitude of pathogens, inducing SAR can eliminate the need for a number of separate agents which would otherwise be necessary to protect a crop, or reduce the amount of the separate agents which would otherwise be required. And, because the induction of SAR can essentially be performed by repetitive action, use of this technique would demand far less effort for the farmer than the currently required regimen of applying multiple agents, each with their own directions for handling, timing, amounts, concentrations, methods of application, and possible adverse interactions.
One of the world""s largest pharmaceutical companies has made an effort to develop the use of systemic inducers to protect crops in the field. To this end, it is bringing to market a systemic inducer, benzothiadiazole, under the trade name Actigard.(trademark) But, the manufacturer now recommends that Actigard(trademark) be used in combination with conventional chemical agents in providing protection to crops. Thus, even a systemic inducer specifically selected, developed and tested for protection of crops has not eliminated the need for conventional pesticides even during the time the systemic inducer is being applied.
U.S. Pat. No. 5,607,856, teaches compositions and methods for sterilizing soil using oxygen radicals. The method involves contacting the soil with a solution of an activated oxygen species, a water-soluble phenolic complex extracted from a material such as humic material, a divalent cation, and a cation redox reducing agent.
What is needed in the art is a means of protecting a variety of crops, flowers, decorative and other plants in the field from pathogens more effectively, at lower cost, and with less effort than by the use of pesticides and other traditional chemical agents. Moreover, what is needed is a means of providing this protection with lower and less lasting damage to the environment than caused by such conventional agents. What is further needed is a means of increasing the protection of crops from pathogens to levels above the levels obtainable by the use of systemic inducers alone, to more crops than can be protected by the use of systemic inducers alone, and against a wider range of pathogens. The present invention addresses these and other needs.
This invention provides novel methods of protecting plants from pathogens. In one group of embodiments, the methods involve contacting the foliage of a plant with a plant systemic inducer and a reactive oxygen species, where the amount of the reactive oxygen species is sufficient to increase the expression of phenylalanine ammonia lyase, glutathione S-transferase, hydroxyproline-rich glycoprotein, chalcone synthase, or pathogenesis-related proteins, in the plant above the level which would be induced by the plant systemic inducer in the absence of the reactive oxygen species. The invention further provides a method of contacting a plant with a plant systemic inducer and a reactive oxygen species, where the amount of the systemic plant inducer is sufficient to increase the expression of phenylalanine ammonia lyase, glutathione S-transferase, hydroxyproline-rich glycoprotein, chalcone synthase, or pathogenesis-related proteins in the plant above the level which would be induced by the reactive oxygen species in the absence of the plant systemic inducer. The increase in pathogenesis-related proteins, phenylalanine ammonia lyase, glutathione S-transferase, or hydroxyproline-rich glycoprotein caused by contacting a plant with both a plant systemic inducer and a reactive oxygen species may be additive compared to the level induced by either the systemic inducer in the absence of the reactive oxygen species or by the reactive oxygen species in the absence of the systemic inducer, or they may be greater than an amount which would be additive.
The plant can be edible by humans or by animals, can be grown for fiber content, such as cotton, can be used for or processed to become a medicine or medicament, or can be for decorative, ornamental, or recreational use, such as turf grass, house plants, flowers, or Christmas trees.
The reactive oxygen species (xe2x80x9cROSxe2x80x9d) can be peracetic acid, hydrogen peroxide, a hydroperoxide, a peroxide, or a phenolic hydroperoxide; ozone is not preferred as an ROS. The plant systemic inducer can be salicylic acid, jasmonic acid, isonicotinic acid, dichloroisonicotinic acid, benzothiadiazole, phosphorous acid, arachidonic acid, or cinnamic acid, can be derived from kelp or other seaweed, or can be a beneficial microbe. The systemic inducer can be humic acid, or can be used in combination with humic acid. The ROS and microbial plant systemic inducer can be administered about 24 hours of one another, whereas an ROS and a chemical plant systemic inducer are preferably administered within one hour of each other and even more preferably are administered together in a mixture, either as, for example, a powder, or, more preferably, a solution. The pathogenesis-related protein can be a product of any of the PR genes having or thought to have a role in protecting plants from pathogens, such as the PR-1, PR-2, PR-3, PR-4 and PR-5 genes. The proteins induced can also be phenylalanine ammonia lyase, chalcone synthase, or a hydroxyproline-rich glycoprotein or other proteins related to strengthening of cell walls or plant defense.
The invention further provides a composition for foliar application to plants comprising a plant systemic inducer and a reactive oxygen species wherein the amount of reactive oxygen species is sufficient to increase the level of a natural plant product selected from the group consisting of: phenylalanine ammonia lyase; hydroxyproline-rich glycoproteins, glutathione S-transferase, chalcone synthase, and pathogenesis-related proteins to a level above the level induced by the plant systemic inducer in the absence of the reactive oxygen species. The composition can further comprise an aqueous solution and detergents, chelating agents or sequestering agents.
In another group of embodiments, the invention provides methods of protecting a plant by contacting one or more roots of a plant with a plant systemic inducer and a reactive oxygen species, where the amount of the reactive oxygen species (ROS) is sufficient to increase the expression of phenylalanine ammonia lyase, glutathione S-transferase, hydroxyproline-rich glycoprotein, chalcone synthase, or pathogenesis-related proteins, in the plant above the level which would be induced by the plant systemic inducer in the absence of the reactive oxygen species, provided that the composition does not comprise an exogenous agent selected from the group comprising a cation redox reducing agent and a divalent cation having redox potential in amounts sufficient to reduce the levels of microorganisms in soil around the roots by 40% or more. The invention further provides a method of contacting one or more roots of a plant with a plant systemic inducer and a reactive oxygen species, where the amount of the systemic plant inducer is sufficient to increase the expression of phenylalanine ammonia lyase, glutathione S-transferase, hydroxyproline-rich glycoprotein, chalcone synthase, or pathogenesis-related proteins in the plant above the level which would be induced by the reactive oxygen species in the absence of the plant systemic inducer, provided that the composition does not comprise an agent selected from the group comprising a cation redox reducing agent and a divalent cation having redox potential in amounts sufficient to reduce the levels of microorganisms in soil around the roots by 40% or more. The systemic inducer can be, or can be mixed with, humic acid.
The invention further provides compositions for soil application to plants comprising a plant systemic inducer and a reactive oxygen species wherein the amount of reactive oxygen species is sufficient to increase the level of a natural plant product selected from the group consisting of: phenylalanine ammonia lyase; hydroxyproline-rich glycoproteins, glutathione S-transferase, and pathogenesis-related proteins to a level above the level induced by the plant systemic inducer in the absence of the reactive oxygen species, provided that the compositions do not comprise an agent selected from the group comprising a cation redox reducing agent and a divalent cation having redox potential in amounts sufficient to reduce the levels of microorganisms in soil around the roots by 40% or more. The systemic inducer can be, or can be mixed with, humic acid.